Self-assembled gels composed of peptide-amphiphile nanofibers have been described as being useful in the templated mineralization of hydroxyapatite. Peptide-amphiphiles enriched with negatively charged amino acids such as phosphoserine and aspartic acid can self assemble into nanofibers and induce hydroxyapatite crystals to grow on the surface of the nanofiber as described by Hartgerink et al., Science, 294, 1683-1688, (2001). In addition to providing sites for hydroxyapatite crystal nucleation, the nanofibers also direct the growth of the hydroxyapatite crystals such that their c-axis is oriented parallel to the long axis of the nanofibers. The ability of the peptide-amphiphile nanofibers to organize and direct the growth of the hydroxyapatite crystals is reminiscent on that observed between collagen fibrils and hydroxyapatite crystals in bone.
The directed growth of hydroxyapatite crystals within organized peptide-amphiphile matrices and scaffolds is an important step toward the regeneration of mineralized materials like bone within the body.
While the preparation of oriented hydroxyapatite crystals on individual or small groups of nanofibers has been demonstrated, scaling the utility of hydroxyapatite or other minerals in bundles of nanofibers or within gels comprising nanofibers maybe limited by non-homogeneous mineralization. Non-homogeneous mineralization of nanofiber bundles or nanofiber gels results in coating of the surface nanofibers of the bundle or gel by the mineral crystals. The formed surface crystals inhibit further diffusion of mineral reagents into the interior of the nanofiber bundle or nanofiber gel and precludes formation of larger homogenous composites. In practical applications such as bone regeneration, it would be desirable that hydroxyapatite crystal growth proceed uniformly throughout the nanofiber gel matrix.
A supramolecular assembly is a material in which the constituent units or building blocks of the assembly are molecules or molecular aggregates. The interaction of the units with each other, usually by non-covalent bonding, determines the final shape and size of the supramolecular assembly. An example of a supramolecular assembly found in biological systems is α-hemolysin which is a seven protein aggregate with a non-symmetric mushroom shape. The α-hemolysin aggregate has a pore or channel that is about 16 Å in diameter, which runs parallel to the aggregate's long axis. The aggressive human pathogen Staphylocuccus aureus uses the asymmetric nature of α-hemolysin to implant its stem into the hydrophobic compartment of cell membranes and the hydrophilic nature of the α-hemolysin's mushroom cap to stabilize it in the extracellular space. It is though α-hemolysin's pore channel that RNA macromolecules from the Staphylocuccus aureus pathogen can invade human cells. Synthetic supramolecular assemblies could be designed and synthesized to mimic the action of α-hemolysin's channel pore for drug delivery or other cell therapies.
The amino acid sequence IKVAV (SEQ ID NO: 1) has been identified in other contexts as important for neuron growth and development. Self assembly of peptide-amphiphiles with the IKVAV (SEQ ID NO: 1) sequence have been reported. These peptide-amphiphiles may facilitate neuron growth and development in supramolecular structures formed by these peptide-amphiphiles. One feature of peptide-amphiphiles having a hydrophobic alkyl tail and the IKVAV (SEQ ID NO: 1) amino acid sequence in the peptide head group is that peptide-amphiphile has more than one amphiphilic moment. The peptide sequence of these and other peptide-amphiphiles can be further modified by covalent attachment of ligands or peptide sequences that can interact with various types of cells. For example, the peptide sequence Arg-Gly-Asp (RGD) occurs in fibronectin and has been found to play an important role in integrin-mediated cell adhesion. Inclusion of the RGD peptide sequence ligand into a suitable peptide-amphiphile is expected to promote cell growth and direct templated mineralization of self assembled supramolecular structures of such peptide-amphiphiles under the proper conditions. Self assembled peptide-amphiphiles are known to direct the mineralization of hydroxyapatite on the surfaces of nanofibers formed from these peptide-amphiphiles. The peptide portion of these peptide-amphiphiles can also comprise amino acid groups like cysteine, which are capable of forming disulfide bonds between adjacent peptide-amphiphiles, and also glycine which provides flexibility to the peptide portion of the molecule.
It will be appreciated by those skilled in the art that there is a need to be able to form self assembled supramolecular structures from peptide-amphiphiles having more than one amphiphilic moment in order to take advantage of the unique cell growth, molecular transport, and templating functions that these and other related peptide sequences provide. It will also be appreciated that the self assembly occur in physiologically benign conditions of temperature, ionic strength, and pH. For the foregoing reasons, there is a need in the art to make supramolecular assemblies from multi-dimensional peptide-amphiphiles.
The present invention is directed to amphiphilic molecular compositions having more than one amphiphilic moment and also to supramolecular composition comprised of such amphiphilic molecules. More specifically, the present invention is directed to peptide-amphiphiles compositions having more than one amphiphilic moment and to supramolecular compositions comprised of such peptide-amphiphiles which self assemble in the presence of cations.
Preferred embodiments of the present invention may be useful for cell growth, molecular transport, and templating functions, especially if the self assembly occurs under benign conditions.
Homogeneously, or substantially homogeneously, mineralized self assembled peptide-amphiphile nanofibers are desirable. Homogenously mineralized materials with the mineral crystals preferentially oriented by the self assembled peptide-amphiphile nanofibers are also desired. Finally, preparing such materials under substantially neutral or physiological conditions is also desirable.